1. Field of the Invention
The present invention relates to a method for preparing sprayable formulations of mycelium-based biological control agents produced by solid state fermentation. More specifically, the present invention relates to a method of production and use of mycelium-colonized particulate substrates in formulations that can be applied with conventional spray equipment. The invention also relates to compositions of sprayable formulations for delivering mycelium-based biological control agents to obtain maximum biological activity and viability.
2. Prior Art
Recent advances in biotechnology have resulted in significant increase in the use of microorganisms as biological control agents in agriculture, forestry, and environmental management. One group of microorganisms that has received particular attention in this area is fungi. A large number of fungi are known for their specific pathogenicity to weeds and insect pests, and many of them have been subjected to thorough scientific studies and commercial development as potential biological control agents. However, very few of these fungi have been commercialized successfully. In many cases, fungal-based biological control agents have failed to reach the market because of the lack of formulations to deliver the products effectively and economically.
As commercial products, biological control agents must be produced and sold in the ways that are more familiar to the end-user: the farmer, forester and environmental engineer. The two main criteria for a commercially viable fungi based product are:
1) that such products must be formulated to preserve maximum cell viability and biological activity under prolonged conditions of shipment and storage; and,
2) that such products must also be formulated and supplied to the end-user in a physical form that does not require new equipment, new technology or new application techniques.
Because the most common and effective application method used today is a spray that is applied with conventional equipment developed for agrochemicals, it is desirable to provide the biological control agents in similar sprayable formulations. For such a formulation, it is important that the biological control agent be provided in a generally uniform size and be dispersible in a liquid carrier so as not to clog sprayer nozzles. In this regard, fungi present a unique problem because of their filamentous structure known as mycelium.
Mycelium is a vegetative form in which a majority of fungal species grow and is very fragile and often varies greatly in sizes and shapes. Although mycelium can be easily produced by fermentation at commercial scale, it has proven difficult to process mycelium into sprayable formulations because of its fragile nature and uneven sizes. This formulation problem has become a major obstacle that blocks many mycelium-based biological control agents from reaching the market and from achieving successful commercialization.
In order to overcome the formulation problem associated with mycelium, several methods and processes have been disclosed in prior art. A simple method is the wet maceration of actively growing mycelium obtained from liquid culture. This method basically involves producing fungal mycelium by submerged fermentation, harvesting of the actively growing mycelium by filtration or centrifugation, reduction of mycelium particle size by high shear blending or milling prior to spray application. This method has been widely used in greenhouse and field experiments. For example, Wall et al. (U.S. Pat. No. 5,587,158) use this method for the application of Chondrostereum purpureum, a biological control fungus for weed trees. Despite its simplicity, the mechanical macerations drastically reduces the viability of the mycelium and often yields a formulation that has very low titer and short shelf life.
In another method, McCabe et al. (U.S. Pat. No. 4,530,834) disclose a dry grinding process for reducing the particle size of mycelium. In this reference, actively growing mycelium harvested from submerged fermentation is dried with protective agents, and then the dry mycelium mass obtained is milled to a form a powder. The dry powder preparation, when it is needed, can be re-wet, diluted in aqueous liquid and applied by spraying. Dried mycelium particles obtained from submerged fermentation of different fungi have been produced in similar ways by other investigators. In such cases, the mycelium obtained from submerged fermentation process is dried, ground in a hammer mill and passed through a sieve to obtain a desired particle size.
Although the dry mycelium powder has extended shelf-life and can be easily stored and handled, it again has a very low titer due to damage caused to the cellular structure of the mycelium by drying and milling.
In short, the prior art, as described above, teaches the use of actively growing mycelium produced by submerged fermentation, which has associated therewith the shortcomings of low titer or poor shelf life. As a result, there is a need for a stable, economical, mycelium-based formulation that can be applied with conventional spray equipment.